1-arabinofuranosyl-5-halouracil and process for the preparation thereof



United States Patent 3,155,646 1 ARAEINUFURANOSYL HALOURACE AND PROCESSFGR THE PREPARATKGN TlEiREtOF James H. Hunter, Kalamazoo, Mich, assignorto The Upjohn Company, Kalamazoo, Mich, a cerporation of Delaware NoDrawing. Filed Aug. 23, 1960, Ser. No. steer 9 Qlairns. (Cl. zen-211.5

This invention relates to novel compounds and to a process for preparingthem. More particularly, the invention is directed tol-arabinofuranosyl-S-halouracil nucleosides and to a process forpreparing them. Still more particularly, the invention relates tol-B-D-arabinofuranosyl-fi-halouracil nucleosides having the followingstructural formula:

0 Hats 5 x wherein X is halogen.

The novel 1-arabinofuranosyl-S-halouracil nucleosides of this inventionare antimetabolites, and they are active against viruses, fungi, andbacteria. The novel 1-arabinofuranosyl-S-halouracil nucieosides can beused for preventing or controlling epizootics of viral and bacterialetiology in mammals and birds, and they are useful as intermediates inthe preparation of the corresponding nucleotides by phosphorylation, forexample, by condensation with phosphoric acid in the presence of yeastenzymes according to the method described in US. Patent 2,844,514. Thel-B-D-arabinofuranosyl 5 halocuracil nucleosides are preferred compoundsof the invention, and they are useful in the treatment of influenza Avirus infections in colonies of laboratory mice, in the treatment or"influenza in swine, in the treatment of Newcastles disease in poultry,in the treatment of canine and feline distemper, and for like diseases.lllustratively, the novel l-fl-D- arab nofuranosyl-S-iodouracil of thisinvention is an effective antivirfl agent; it is active againstinfluenza A (PR- 8) and Western equine encephalomyelitis viruses.

The novel l-arabinofuranosyl-5-halouracil nucleosides of this inventionare prepared by direct halogenation of a l-arabinofuranosyluracil.Illustratively, l-arabinofuranosyl-5-chloro-,1-arabinofuranosyl-5-bromo, and l-arabinofuranosyl 5 iodouracil areprepared by direct halogenation, advantageously, by reactingl-arabinofuranosyluracil with, respectively, chlorine, bromine, andiodine in the presence of an inert solvent which can be aqueous ornonaqueous. In general, selection of the inert solvent depends on theparticular halogen, as will be readily apparent to those skilled in theart. Illustratively, chlorination can be carried out advantageously inthe presence or" a non-aqueous solvent, for example, a mixture of aceticacid and carbon tetrachloride; bromination in the presence of Wateralone; and iodination in the presence of water, an organic solvent, forexample, chloroform, and an oxidizing agent, for example, nitric acid.If desired, the reaction can be accelerated by gently heating thereaction mixture. The l-arabinofuranosyl-5-halouracil thus produced isthen recovered by conventional methods, e.g., by removing excesshalogen, evaporating the volatile components of the reaction mixture,solvent extraction, and crystallization; and like procedures.

If desired, direct halogenation can be carried out with apoly-O-acylated l-arabinofuranosyluracil, for example,1-(2,3,5-tri-O-acetyl-,B-D-arabinofuranosyl)uracil and the like. Theresulting poly-O-acylated l-arabinofuranosyl- S-halouracil is thendeacylated by hydrolysis, advantageously by acid hydrolysis, in order toobtain the desired 1-arabinoiuranosyl-S-halouracil. When this procedureis used, the hydroxyl groups of the l-arabinofuranosyluracil are blockedwith the acyl group of a monocarboxylic acid, advantageously, ahydrocarbon carboxylic acid containing from 2 to 8 carbon atoms,inclusive. Acylation of a l-arabinofuranosyluracil is accomplished byreaction With the acid anhydride or the acid chloride of amonocarboxylic acid such as, for example, acetic anhydride, acetylchloride, or benzoyl chloride, and like acylating agents, advantageouslyin the presence of an acid acceptor, for example, pyridine, picoline,collidine, triethylamine, and the like.

Suitable such acids include saturated and unsaturated straight orbranched chain aliphatic carboxylic acids such as acetic, propiom'c,butyric, isobutyric, tert-butyl-acetic, valeric, isovalerie, caproic,caprylic, and the like; and aromatic carboxylic acids such as benzoic,toluic, ethylbenzoic, phenylacetic and the like.

The lfi-D-arabinofuranosyl-S-halouracil nucleosides of this inventioncan also be prepared, advantageously, by

preparing a 1-,8-D-ribofuranosyl-S-halouracii and then enemas 1.23quence of reaction steps and intermediates prepared during theepimerization process are as follows:

The 1 arabinofuranosyl 5 halouracil nucleosides of this invention formsalts with bases, and it will be understood that this invention includessalts obtained by reacting a l-arabinofuranosyl 5 -halouracil withbases, e.g., alkali metal hydroxides, alkaline earth metal' hydroxides,ammonia, non-toxic organic bases such as ethanolamine, and the like, toform pharmacologically acceptable salts.

The following examples are illustrative of the process and products ofthe present invention, but are not to be construed as limiting.

EXAMPLE 1.PREPARATION OF l-fl-D-ARAB- INOFURANOSYL-S-CHLOROURACIL Asolution of 370 mg. (1.0 milljmole) of 1-(2,3,5-tri-O-acetyl-fl-D-arabinofuranosyl)uracil in ml. of glacial acetic acid wasmixed with 5 ml. of a 1.56% (w./v.) solution of chlorine in carbontetrachloride (the 5 ml. contained about 78 mg. or 1.1 millimoles ofchlorine). The reaction mixture was held at about 25 C. for 34 hrs.(external moisture was excluded). The volatile components were thenremoved by distillation at about 25 C. and 0.2 to 1.5 mm. of mercurypressure, and the material (a glass) thus obtained was held at 15 to mm.of mercury pressure and about C. for about 14 hrs. The glass wasdissolved in warm absolute ethanol, and the ethanol was removed bycodistillation (initially at a temperature of about 40 C. and 15 to 20mm. of mercury pressure, and finally at a temperature of 40 to 55 C. and0.2 to 0.3 mm. of mercury pressure). This codistillation with absoluteethanol was repeated. The amorphous 1 (2,3,5 tri O acetyl {3 Darabinofuranosyl) 5- chlorouracil thus obtained was dissolved in 10 ml.of absolute methanol and 200 mg. of dry hydrogen chloride was added. Thereaction vessel was stoppered securely, and it was held at about 25 C.for 18.5 hrs. The volatile components were partially removed bydistillation at about 25 C. and 15 to 20 mm. of mercury pressure, anddistillation was completed after raising the temperature to about 40 C.The white crystalline material thus obtained was held under reducedpressure at a bath temperature of about 40 C., and it was then powderedand triturated with acetone. The resulting white solid was collected ona filter; the filter cake was washed with acetone and dried in air.There was thus obtained 220 mg. of1-,8-D-arabinofuranosyl-S-chlorouracil having a melting point of 224 to226 C. The product was dissolved in hot water and the aqueous solutionwas filtered through a bed of acid-washed filter aid (Celite). Thefilter was Washed with hot water, and the combined filtrate and washingswere allowed to cool. As the solution cooled, large rosettes of whiteneedles began to form. The crystallizing mixture was held at about 25 C.for about 8 hrs. and refrigerated at 0 C. for about 12 hrs. The crystalsof 1-B-D-arabinofuranosyl-5-chlorouracil were collected on a filter,washed with ice-water and dried in air. They weighed 218 mg. (78.5%yield) and had a melting point of 231 to 232 C.

Analysis:

Calculated for C H ClN O C, 38.79; H, 3.98; Cl,

12.72; N, 10.06. Founcl-C, 38.84; H, 13.93; C], 12.74; N, 10.23.

Optical rotation:

[a] +77 (c., 0.730 water).

Ultraviolet absorption:

A355, m 239 mu (A 1,345) Nf ifif 276 III/4 (A 9,446)

moi; 24 my (A 3,147) x1353? 275-276 mu Characteristic infraredabsorption frequencies (cm- OH/NH 3340, 3270, 3150.

Unsat. CH 3040, 3020.

C-O/C=C 1715, 1702, 1680, 1620. C O/C- -N 1283, 1205, 1165, 1118, 1085,

1046, 1032, 1017. Armo. sub. 800, 680.

R; 0.62 water-saturated n butanol, descending), 0.83 (v./ v.) aqueousisopropanol 2 N HCl, descending).

EXAMPLE 2.-PREPARATION OF l-fi-D-ARABINO- FURANOSYL-S-BROMOURACIL Aquantity (448 mg., 1.8 millimoles) of finely dividedl-fl-D-arabinofuranosyluracil was dissolved in 4 ml. of saturatedbromine water. To the resulting clear solution was added 11 ml. ofbromine water in portions of 1 ml. each. The reaction vessel was swirledafter each addition. The reaction mixture which became yellow afteradding the last portion of bromine water, was then swirledintermittently for 5 to 10 minutes; it was then decolorized by bubblingthrough it a slow stream of nitrogen gas. The colorless solution waschilled and concentrated to approximately one-half its original volumefrom a semi-frozen state under reduced pressure; it was then stored forabout 14 hrs. at 15 to 20 mm. of mercury pressure. The volatilecomponents were removed at about 30 C. and 0.2 to 0.5 mm. of mercurypressure, and the residue was mixed with 15 ml. of absolute ethanol andrefluxed for approximately 15 minutes. The mixture was cooled and thenrefrigerated at -20 C. A crystalline solid that separated was collectedon a filter, washed with cold absolute ethanol and dried in air. Thecrude l-fi-D-arabinofuranosyl-S-bromouracil thus obtained weighed 440mg; it was dissolved in hot 95% ethanol and the solution was Sitrredwith decolorizing charcoal. The suspension was filtered, and 200 mg. ofcrystalline l-fi-D-arabinofuranosyl-S-brornouracil was obtained from thefiltrate.

Analysis:

Calculated for C H B1N O C, 33.45; H, 3.43; Br,

24.73; N, 8.67. Found: C, 33.59; H, 3.55; Br, 23.95; N, 8.53. Opticalrotation:

[a] +49 (c., 0.400 water). Ultraviolet absorption:

A9 1,, 243 mu (A 1,459) 19, 13 27827 9 mp A251,? 250 my. (A 2,713)Ail-$5 277-278 mu Characteristic infrared absorption frequencies (cmfOPT/NH 3340, 3280, 3210, 3160. Unsat. CH 3050, 3000. C O/C -N 1715 sh.,1700, 1675, 1667. @C 1613. C-O/CN 1308, 1282, 1080, 1073,

1050, 1030, 1014. Arom. sub 855, 798, 772, 740, 680.

R 0.51 (water-saturated n-butanol, descending), 0.86 (65% (v./v.)aqueous isopropanol-Z N HCl,

descending) EXAMPLE 3.-PREPARATION OF l-fi-D-ARABINO-FURANOSYL-S-IODOURACIL A mixture consisting of 448 mg. (1.8 millimoles)or" 1-,8-D-arabinofuranosyluracil, 456 mg. (1.8 millimoles) of iodine,2.5 ml. of chloroform, and 4.9 ml. of 1.0 N nitric acid was heated undergentle reflux for about 5 hrs. The reaction mixture was cooled, and thenrefrigerated at 0 C. The solids which separated were collected on afilter, washed extensivelywith ether, and dried in air. The crudeproduct thus obtained weighed 560 mg. and had a melting point of 190 to195 C., with previous softening. This crude product was dissolved in hotwater, decolorizing charcoal was added, and the suspension was stirredand filtered. Crystallization occurred spontaneously, and was completedby refrigerating at 0 C. for about 18 hrs. The crystals of purifiedl-fi-D- arabinofuranosyl-S-iodouracil were collected on a filter, washedwith ice-water and dried in air. They weighed 315 mg. and had a meltingpoint of 196 to 199 C.

Analysis:

Calculated for C H IN O -C, 29.20; H, 3.00; I,

34.29; N, 7.57. Found--C, 29.21; H, 2.80; I, 34.05; N, 7.56. Opticalrotation:

[a] +30 (0., 0.7396 water). Ultraviolet absorption:

khin 246-248 111;].(A 1,801) A851,? 289-290 mu Afilfl 253 m (A 2,225) A31,}? 27 9-283 ma Characteristic infrared absorption frequencies (cm.-OH/NH 3400 sh., 3310, 3220 sh.,

3060. C=O 1685, 1675, 1663. C C 16 07, 1480 sh. C-O 1150, 1135, 1075sh., 1065,

6 Atom. sub 875, 808, 790, 710.

R 0.51 (water-saturated n-butanol, descending), 0.85 (65% (v./v.)aqueous isopropanol-2 N HCl, descending).

EXAMPLE 4.-PREPARATION OF l-u-D-ARABINO- FURANOSYL-S-IODOURACILFollowing the procedure of Example 3, but substituting1wt-D-arabinofuranosyluracil for l-B-D-arabinofuranosyluracil;1-a-D-arabinofuranosyl-S-iodouracil was prepared.

EXAMPLE 5.-PREPARATION OF l-[i-D-ARABINO- FURANOSYL-S-FLUOROURACIL PartA.--Preparati0n 0f 1-(2,3-O-Isopr0pylidene fi-D- Ribon furanosyl 5 -Fluorouracil A mixture consisting of 26.2 g. (0.1 mole) of l-fi-D-ribofuranosyl-S-fiuorouracil, 49 g. of anhydrous cupric sulfate, 600 ml.of acetone, and 0.65 ml. of concentrated sulfuric acid was stirred andmaintained at approximately 25 C., for 70 hrs. (external moisture wasexcluded). The reaction mixture was filtered and the filter cake waswashed with acetone. The filtrate and washings were combined and 25 g.of calcium hydroxide was added. After stirring the suspension for 3 hrs,it was filtered, and the filter cake was washed repeatedly with acetone.After combining the filtrate and washings, the volatile components wereevaporated, and 1-(2,3-O-isopropylidine-fi-D-ribofuranosyl)-5-fluorouracil was recovered.

Part B.-Preparati0u 0f1-(S-O-Acezyl-2,3-O-Isopr0pylidine-B-D-Rib0furan0syl) -5-Fluor0uracil Asolution of 1-(2,3-0-isopropylidine-fl-D-ribofuranosyl)-5-fluorouracil(prepared in Part A) in pyridine was treated with an excess of aceticanhydride. The reaction mixture was held at 20 to 25 C. for about 5 hrs.The mixture was cooled, methanol was added, and the volatile componentswere removed under reduced pressure at a bath temperature of about 50 C.The residual solid thus obtained was dissolved in absolute ethanol andthe ethanol was removed under reduced pressure at a bath temperature ofabout 50 C. This codistillation procedure was repeated two times and theresulting 1-(5-0- acetyl 2,3 O isopropylidine-B-D-ribofuranosyl)-5-fiuorouracil was purified by crystallization from 50% (v./v.) aqueousethanol.

Part C.Preparati0n of Z-(S-O-Acetyl-fl-D-Ribofuranosyl -5-Flu0rouracilThe1-(5-O-acetyl-2,3-O-isopropylidine-fl-D-ribofuranosyl)-5-iluorouracil(prepared in part B) was suspended in approximately 20 volumes of 70%(v./v.) aqueous acetic acid, and the suspension was heated at about 75C. for about 5 hrs. The volatile components were then removed at 0.3 to0.7 mm. of mercury pressure and a bath temperature of 70 C. The syrupyresidue was dissolved in absolute ethanol, and the ethanol was removedunder reduced pressure and a bath temperature about 70 C. Thiscodistillation procedure was repeated, and the residual crude1-(5-0-acetyl-B-D-ribofuranosyl) -5-fiuorouracil was purified bycrystallization from ethanol.

Part D.Preparati0n 0f 1-(S-O-AcetyZ-Z-O-Tosyl-(i-D- Ribofuranosyl-5-Flu0r0uracil A solution of 3.04 g. (10 millimoles) of 1-(5-O-Acetyl-B-D-ribofuranosyl)-5-fiuorouracil (prepared in Part C) in 20 ml. ofpyridine was mixed with 1.9 g. (10 millirnoles) of p-toluenesulfonylchloride. The reaction vessel was stoppered securely, and held at 20 to25 C. for 15 hrs. The mixture was chilled and methanol was added. Thevolatile components were removed at about 60 C. (initially at about 20mm. of mercury pressure, and finally at about 0.5 mm. of mercurypressure). The material thus areas re obtained was swirled with warmabsolute ethanol and the ethanol was removed under reduced pressure at abath temperature of 60 to 70 C. After trituration with warm absoluteethanol and refrigeration at C., there was obtained1-(5-O-acetyl-2-0-tosyl-fl-D-ribofuranosyl)-5-fiuorouracil.

Part E .Prcparali0n 0 f I -O-A cetyl-O 2-CycIo-@-D- A rabz'nofuranosyl-5-Flu0r0umcz'l A mixture of1-(S-O-acetyl-2-O-tosyl-fi-D-ribofuranosyl)-5-fiuorouracil (prepared inPart D) and an excess of sodium azide in 95% ethanol was heated underreflux for 6 hrs. in a stream of nitrogen (the escaping gases werebubbled through aqueous sodium hydroxide). The volatile components werethen removed under reduced pressure at a temperature of about 70 C. Thematerial thus obtained was triturated with hot acetonitrile and theacetonitrile was then removed at a bath temperature of about 70 C.(initially under 15 to 20 mm. of mercury pressure and finally under 0.5mm. of mercury pressure). The l- S-O-acetyl-O,2-cyclo-/9D-arabinofuranosyl -5-1'iuorouracil was separated from anysodium p-toluenesulfonate by repeated extraction with boilingacetonitrile and evaporation of the acetonitrile.

Par! F .-Preparalion of J-fl-D-Arabinofumzzosyl-5- F luorouracil Aquantity (5.72 g., 0.02 mole) of l-(5-O-acetyl-O ,2-cyclo-fl-D-arabinofuranpsyl)-S-fluorouracil was dissolved in 0.1 Nsulfuric acid and the reaction mixture was heated on a steam bath for 3hrs. The reaction mixture Was cooled and made faintly alkaline tophenolphthalein with 0.1 N barium hydroxide. The precipitated bariumsulfate was separated by centrifugation and the clear supernatant wasadjusted to about pH 5 with a stream of carbon dioxide gas. The somewhatcloudy solution thus produced was filtered and the filtrate concentratedin a stream of air to about two-thirds of the original volume. Thethusprecipitated barium carbonate was separated by filtration and thefiltrate was evaporated to dryness at 0.3 mm. of mercury pressure and abath temperature of to C. After grinding the resulting solid to a powderand drying under reduced pressure over phosphorous pentoxide, crystalsof l-,8-D-arabinofuranosyl-5-fluorouracil were obtained by dissolving inhot methanol, concentrating the solution, and chilling.

I claim:

1. Compound selected from the group consisting oflarabinofuranosyl-S-halouracil and pharmacologically acceptable saltsthereof.

2. Compound selected from the group consisting of 1-18-D-arabinofuranosyl-S-halouracil and pharmacologically acceptable saltsthereof.

3. 1-,8D-arabinofuranosyl-5-halouracil wherein halo is a halogen havingatomic weight between 35 and 127, inclusive.

4. l-fi-D-arabinofuranosyl-S-chlorouracil.

5. l-fi-D-arabinofuranosyl-S-bromouracil.

6. 1-5-D-arabinofuranosyl-S-iodouracil.

7. The process for preparing 1-arabinofuranosyl-S-halouracil whichcomprises reacting l-arabinofuranosyluracil with a halogen in thepresence of a solvent and recovering l-arabinofuranosyl-S-halouracil.

8. The process for preparing l-,B-D-arabinofuranosyl-5- haiouracil whichcomprises reacting l-p-D-arabinofuranosyluracil with a halogen in thepresence of a solvent and recoveringl-fi-D-arabinofuranosyl-S-halouracil.

9. The process according to claim 8 wherein halo is a halogen having anatomic weight between 35 and 127, inclusive.

OTHER REFERENCES Brown et al., Jr. of Chem. Soc. (London), July 1956,pp. 23882393.

UNITED STATES PATENT OFFICE CERTIFICATE OF CORRECTION Patent No. 3,155,646 November 3, 1964 James H. Hunter It is hereby certified that.error appears in the above numbered patent requiring correction and thatthe said Letters Patent should read as corrected below.

Column 4, line 44, for C:O/C=N" read C-O/C-N Signed and sealed this 30thday of March 1965,

( SEA L) Attest:

EDWARD J. BRENNER Commissioner of Patents ERNEST W. SWIDER AitcstingOfficer

1. COMPOUND SELECTED FROM THE GROUP CONSISTING OF1ARABINOFURANOSYL-5-HALOURACIL AND PHARMACOLOGICALLY ACCEPTABLE SALTSTHEREOF.